The sequence listing filed herewith, titled “Sequence_Listing,” having a file size of about 5,196 bytes and created on Jul. 1, 2011 is hereby incorporated by reference in its entirety.
The present invention relates to penaeidin gene promoters found in tiger shrimps and applications thereof.
Tiger shrimps are the most economically important shrimp species in the world. Large-scale tiger shrimp cultures exist in Southeastern Asia, South Pacific, and Middle and South America, which generated over 400,000 tons of production and over 3 billion U.S. dollars of revenue in 1997. However, the tiger shrimp culture industry in Taiwan almost collapsed in recent years because of the great loss caused by the deterioration of culture environment and the outburst of infectious diseases.
To save tiger shrimp culture industry in Taiwan, many studies aimed at the aspects of culture environment, feed, shrimp diseases, selection of specific-pathogen-free (SPF) shrimp spawners or fry, and shrimp breeding, for example. However, it is difficult to achieve the purpose of disease control by altering the culture environment, such as adjusting the temperature, salt level, pH value, and NH3 level since the optimal growing conditions of pathogens usually overlap with those of shrimps. In addition, although the survival rate of shrimps can be raised by supplementing feed with glucan that increases the general disease resistance, it is limited by the narrow dose range of immunostimulants, and negative effects are resulted if high doses are used. Furthermore, although the technique for selecting SPF shrimp spawner or fry has matured, it is still costly to screen for the shrimp spawner or fry that contains no commonly known pathogens, and the number of such shrimp spawner or fry is small currently. Besides, the thus obtained SPF shrimps must be farmed in isolated space with circulating water. If they are farmed in a traditional outdoor pool where it is hard to keep these shrimps away from pathogens, their mortality rate would even exceed that of shrimps with a slight illness. As for the use of antibiotics, there exists the problem of residue although they are effective. The prices of shrimps farmed in China, Thailand, Vietnam and India have fallen greatly since the European Union published the report on antibiotic residue in farmed shrimps, seriously damaging the development of shrimp farmers industry.
Penaeidins belong to a family of antimicrobial peptides that exhibit both Gram-positive antibacterial and antifungal activities. They have been found in Litopenaeus, Marsupenaeus, Fenneropenaeus, Penaeus and Farfantepenaeus shrimps. Penaeidins are constitutively synthesized and stored in the shrimp hemocytes, located in granulocyte-cytoplasmic granules, and released in response to appropriate stimuli such as infections. Three classes of penaeidins, PEN2, PEN3 and PEN4, were identified in the Pacific white shrimp L. vannamei, each class being encoded by a unique gene, and among which PEN3 exhibited the highest mRNA expression. Tiger shrimp Penaeus monodon was found to have one class of penaeidin, high mRNA expression of which was detected during the nauplius I and intermoult stages.
There are many successful examples of introducing heterologous genes which provide desired characteristics of strong disease-resistance, high stress-endurance, and increased growth rate into fish and shellfish by genetic transformation techniques. Tseng et al. introduced heterologous DNA into tiger shrimps by electroporation to express a reporter gene (Theriogenology. 2000; 54(9):1421-32). They used an expression vector comprising a promoter derived from a mammalian virus. Although the promoter can be recognized by most biological systems, the disadvantage is leaving a virus-derived DNA fragment in the transgenic shrimp.